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Indirect fluorescent antibody staining

Whereas multicolor immunoenzyme staining is applicable only for separately located antigens (see Chap. 7), multicolor fluorescence immmunostaining makes it possible to colocalize antigens not only in the same cell but also in the same cellular compartment. Simultaneous immunolocalization of antigens using fluorescent antibodies can be fulfilled both by the direct (see Sect. 4.1) and indirect (see Sect. 4.2) methods. With the direct method, primary antibodies are labeled with fluorescent dyes, while with the indirect method, primary antibodies are applied as unlabeled antibodies and the visualization is performed with secondary antibodies that are labeled with fluorescent dyes. [Pg.69]

The actual response of monoclonal antibodies with individual cells is usually visualized either directly (typically using fluorescent stains) or indirectly [using the reaction of antibody labeled with horseradish peroxidase (HRP) or other enzymes] with diaminobenzidine (DAB) (or other substrate while using other enzymes) under the microscope or in the flow cytometer. The latter, however, is not employed routinely in CSF immunocytology, although it has an advantage in clinical hematology. [Pg.55]

Triton X-100). Alternatively, cells can be fixed and permeablized with organic solvents such as methanol and acetone. The type of fixative depends on the cellular components that need to be detected and the employed antibodies. Cellular components are then detected either directly by means of dyes (e.g., fluorescent dyes that bind to and stain DNA) or antibodies that are labeled with fluorophores. Alternatively, an indirect detection can be performed by means of primary antibody that specifically binds to a protein of interest and a secondary antibody that recognizes the primary antibody and is labeled for detection (usually with a fluorophore). Finally, samples are analyzed by means of fluorescence microscopy. [Pg.240]

Changes indnced by mechanical stimnlation on the cy toskeleton were qualitatively assessed through indirect immunofluorescence. Primary antibodies against actin, laminin and tubulin were used stains show stronger fluorescence on mechanical stimulated cells, clearer images of the cytoskeleton elements and nucleus delimitation and prominent cytoplasmatic extensions (Figures 2 to 4). [Pg.296]


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See also in sourсe #XX -- [ Pg.531 ]




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